Viral and non-viral vectors are used as the basis for gene delivery in current nucleic acid and gene therapy methods. However, there are concerns about the production, reproducibility, cost, and safety of viral vectors for gene therapy. As a result, work has focused on the development of nonviral vectors where the gene construct of interest is delivered by synthetic nonviral materials. Examples of such materials include polycations, dendrimers, and polysaccharides, as well as small molecule cationic lipids such 3-beta-[N′,N′dimethylaminoethane)-carbamoyl]cholesterol (DC-chol). Cationic lipids for lipofection condense plasmids, facilitate enclose escape, neutralize charge of DNA, and/or shield DNA from nucleases (basic, 1997, Liposomes in Gene Delivery, CRC Press).
Cationic lipids facilitate plasmid delivery and some cationic sterol-based compounds are known to possess antimicrobial activity due to their amphiphilic character. Given the persistent bacterial infection associated with several diseases targeted by gene therapy such as cystic fibrosis (Beadier, 2007, Annual Rev Med, 58:157-170) and the potential consequence of infections on the efficacy of gene delivery administration, antibacterial activity exhibited by the gene delivery vehicle would offer a therapeutic benefit.
Recently, several novel steroidal dimers have shown activity against certain pathogens and some compounds have been used to facilitate both in vitro transfection and bactericidal activity. (Blagbrough et al., 2003, Biochem Soc Trans, 31:397-406; Kichler et al., 2005, J Control Release, 107:174-182; Salunke et al., 2004, J Med Chem, 47:1591-1594). Facially amphiphilic lipid structures are believed to interact with membranes by an analogous mechanism to naturally incurring peptide antibiotics which are active against both gram-positive and gram-negative bacteria.
Cationic lipids are commonly used non-viral vectors for gene delivery duo to their ability to condense plasmid DNA (Hirko et al., Curr Med Chem, 10:1185-1193). Following synthesis of N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA) for lipofection (Felgner et al., 1987, PNAS, 84:7413-7417), optimization of the molecular structures of cationic lipids has been an active area of research including headgroup (Narang et al., 2005, Bioconjug Chem, 16:156-168; Obata et al., 2008, Bioconjug Chem, 19:1055-1063), linker (Bajaj et al., 2008, J Med Chem, 51: 2533-2540; Rajesh et al., 2007, J Am Chem See, 129:11408-11420; Aissaoui et al., 2004, J Med Chem, 47:5210-5223), and hydrophobic domain modifications (Remy et al., 1994, 5:647-654; Heyes et al., 2002, J Med Chem, 45; 99-114). Important modifications have included the use of multivalent polyamines, (Behr et al., 1989, PNAS, 86:6982-6986) which improve DNA binding and delivery via enhanced surface charge density (Martin et al., 2005, Curr Pharm Des, 11:375-394) and the use of sterol-based hydrophobic groups such as 3B—[N—(N,N-dimethylaminoethane)-carbamoyl]cholesterol (DC-Chol), which limits toxicity (Gao and Huang, 1991, Biochem Biophys Res Commun, 179:280-285). Helper lipids such as dioleoyl phosphatidylethanolamine (DOPE) are used to improve transgene expression via enhanced liposomal hydrophobicity and hexagonal inverted phase transition to facilitate endosomal escape (Karanth et al., 2007, J Pharm Pharmacol, 59:469-483). Studies of mixed lipids are less common; however, recent studies involving mixtures of cationic lipid derivatives have shown promise and represent an interesting new area for optimization (Wang and MacDonald, 2004, Gene Ther, 11:1358-1362; Wang and MacDonald, 2007, Mol Pharm, 4:615-623; Caracciolo et al., 2007, Biochim Biophys Acta, 1768:2280-2292). In addition to the molecular structures of cationic lipids, transfection efficiency has been linked to physicochemical characteristics and morphology of structures formed following complex formation with DNA (Ma et al., 2007, J Control Release, 123:184-194). Critical factors influencing transfection activity include lipoplex charge ratio (lipid:DNA), solution ionic strength, and residual net surface charge of lipoplexes (liposome-DNA complex).
Interestingly, several findings have indicated that inflammatory cytokines can inhibit gene transfer in vitro with a decrease in both transcription and transgene activity of ˜50%. (Baatz et al., 2001, Biochim Biophys Acta, 1535:100-109; Bastonero et al., 2005, J Gene Med, 7:1439-1449). This inhibitory effect was prevented by glucocorticoid treatment indicating blocking the NF-KB pathway, which is known to control upregulation of numerous inflammatory cytokines including IL-8 and TNF-alpha (Kulms and Schwarz, 2006, Vitam Horm, 74:283-300), may play a critical role between induced inflammation and efficiency of gene transfer. In addition to the pathogenesis associated with infection, bacterial membrane bound molecules, such as lipopolysaccharide (LPS), are known to activate a strong inflammatory response in eukaryotic cells via toll-like receptors (TRL), especially TRL4, (Schnare et al., 2006, Int Arch Allergy Immunol, 139:75-85) therefore, prevention of bacterial-mediated inflammation may also have a direct impact on gene delivery efficiency.
The activities of two sterol-based cationic lipids: dexamethasone-spermine (DS), (Gruneich et al., 2004, Gene Ther, 11:668-674) and disubstituted spermine (D2S), resulting from conjugation of dexamethasone to the polyamine spermine DS, has been shown to exhibit anti-inflammatory activity in an in vivo mouse intraperitoneal thioglycollate challenge model based on neutrophil infiltration and has been shown to condense and deliver plasmid DNA enabling in vitro transfection of plasmid DNA. DS has also been shown to improve airway targeting, attenuate vector-induced inflammation, and facilitate re-administration in vivo when formulated with adenovirus vectors, (Price et al., 2005, Mol Ther, 12:502-509; Price et al., 2007, Gene Ther, 14:1594-1604; see also U.S. patent application Ser. No. 12/259,097).
There is a long felt need in the art for new antimicrobial compositions and methods of treatment. The present invention satisfies this need.